Expression of Membrane Proteins at the Escherichia coli Membrane for Structural Studies
Structural biology of membrane proteins is often limited by the first steps in obtaining sufficient yields of proteins because native sources are seldom. Heterologous systems like bacteria are then commonly employed for membrane protein over-expression. Escherichia coli is the main bacterial host used. However, overproduction of a foreign membrane protein at a non-physiological level is usually toxic for cells or leads to inclusion body formation. Those effects can be reduced by optimizing the cell growth conditions, choosing the suitable bacterial strain and expression vector, and finally co-expressing the target protein and the b-subunit of E. coli adenosine triphosphate (ATP)-synthase, which triggers the proliferation of intracytoplasmic membranes. This chapter is devoted to help the experimenter in choosing the appropriate plasmid/bacterial host combination for optimizing the amount of the target membrane protein produced in its correct folded state.
Key wordsBacterial mutant b-subunit of F-ATPase co-expression internal membrane
This work was supported by INSERM, CNRS and the “Agence Nationale de la Recherche” (grant 05-JCJC-0092-01 to B.M.).
- 4.Lundberg U, Kaberdin V, von Gabain A (1999) The mechanism of mRNA degradation in bacteria and their implication for stabilization of heterologous transcripts. In: Demain AL, Davies JE (eds) Manual of industrial microbiology and biotechnology. ASM Press, Washington, DC, pp 585–596Google Scholar
- 13.Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
- 14.Walker JE, Miroux B (1999) Selection of Escherichia coli host that are optimized for the overexpression of proteins. In: Demain AL, Davies JE (eds) Manual of industrial microbiology and biotechnology. ASM Press, Washington, DC, pp 575–584Google Scholar